chr13-32326240-A-G

Position:

• chr13-32326240-A-G

Variant summary

Our verdict is Pathogenic. Variant got 18 ACMG points: 18P and 0B. PVS1 PM2 PP5_Very_Strong

The NM_000059.4(BRCA2):​c.476-2A>G variant causes a splice acceptor change involving the alteration of a non-conserved nucleotide. The variant was absent in control chromosomes in GnomAD project. In-silico tool predicts a benign outcome for this variant. 3/3 splice prediction tools predicting alterations to normal splicing. Variant has been reported in ClinVar as Pathogenic (★★★).

• Frequency: Genomes: not found (cov: 33)
• Consequence: BRCA2 NM_000059.4 splice_acceptor
• Scores: Splicing: ADA: 1.000
• Clinical Significance: Pathogenic reviewed by expert panel P:13
• Conservation: PhyloP100: 3.54

Genes affected

BRCA2 (HGNC:1101): (BRCA2 DNA repair associated) Inherited mutations in BRCA1 and this gene, BRCA2, confer increased lifetime risk of developing breast or ovarian cancer. Both BRCA1 and BRCA2 are involved in maintenance of genome stability, specifically the homologous recombination pathway for double-strand DNA repair. The largest exon in both genes is exon 11, which harbors the most important and frequent mutations in breast cancer patients. The BRCA2 gene was found on chromosome 13q12.3 in human. The BRCA2 protein contains several copies of a 70 aa motif called the BRC motif, and these motifs mediate binding to the RAD51 recombinase which functions in DNA repair. BRCA2 is considered a tumor suppressor gene, as tumors with BRCA2 mutations generally exhibit loss of heterozygosity (LOH) of the wild-type allele. [provided by RefSeq, May 2020]

ACMG classification

Verdict is Pathogenic. Variant got 18 ACMG points.

• PVS1: Splicing +-2 bp (donor or acceptor) variant, LoF is a know mechanism of disease, No cryptic splice site detected. Exon removal results in frameshift change.
• PM2: Very rare variant in population databases, with high coverage
• PP5: Variant 13-32326240-A-G is Pathogenic according to our data. Variant chr13-32326240-A-G is described in ClinVar as [Pathogenic]. Clinvar id is 37923.Status of the report is reviewed_by_expert_panel, 3 stars. Variant chr13-32326240-A-G is described in Lovd as [Likely_pathogenic]. Variant chr13-32326240-A-G is described in Lovd as [Pathogenic].

Transcripts

RefSeq

Gene	Transcript	HGVSc	HGVSp	Effect	#exon/exons	MANE	Protein	UniProt
BRCA2	NM_000059.4	c.476-2A>G		splice_acceptor_variant		ENST00000380152.8	NP_000050.3

Ensembl

Gene	Transcript	HGVSc	HGVSp	Effect	#exon/exons	TSL	MANE	Protein	Appris	UniProt
BRCA2	ENST00000380152.8	c.476-2A>G		splice_acceptor_variant		5	NM_000059.4	ENSP00000369497	A2

Frequencies

GnomAD3 genomes Cov.: 33

GnomAD4 exome Cov.: 31

GnomAD4 genome Cov.: 33

Alfa AF: 0.000298 Hom.: 0

ClinVar

Significance: Pathogenic

Submissions summary: Pathogenic:13

Revision: reviewed by expert panel

Submissions by phenotype

Breast-ovarian cancer, familial, susceptibility to, 2 Pathogenic:5

Pathogenic, no assertion criteria provided	clinical testing	Sharing Clinical Reports Project (SCRP)	May 01, 2012	- -
Pathogenic, criteria provided, single submitter	clinical testing	Consortium of Investigators of Modifiers of BRCA1/2 (CIMBA), c/o University of Cambridge	Oct 02, 2015	- -
Pathogenic, no assertion criteria provided	clinical testing	Breast Cancer Information Core (BIC) (BRCA2)	Feb 20, 2004	- -
Pathogenic, reviewed by expert panel	curation	Evidence-based Network for the Interpretation of Germline Mutant Alleles (ENIGMA)	Aug 10, 2015	IARC class based on posterior probability from multifactorial likelihood analysis, thresholds for class as per Plon et al. 2008 (PMID: 18951446). Class 5 based on posterior probability = 1 -
Pathogenic, criteria provided, single submitter	clinical testing	Counsyl	Jul 14, 2017	- -

not provided Pathogenic:3

Pathogenic, criteria provided, single submitter	clinical testing	Revvity Omics, Revvity	Dec 14, 2022	- -
Pathogenic, criteria provided, single submitter	clinical testing	Quest Diagnostics Nichols Institute San Juan Capistrano	Jun 01, 2017	- -
Pathogenic, criteria provided, single submitter	clinical testing	Mayo Clinic Laboratories, Mayo Clinic	Apr 20, 2023	PP5, PM2, PVS1 -

Hereditary breast ovarian cancer syndrome Pathogenic:2

Pathogenic, criteria provided, single submitter	clinical testing	Labcorp Genetics (formerly Invitae), Labcorp	Sep 21, 2023	This sequence change affects an acceptor splice site in intron 5 of the BRCA2 gene. RNA analysis indicates that disruption of this splice site induces altered splicing and may result in an absent or disrupted protein product. This variant is not present in population databases (gnomAD no frequency). Disruption of this splice site has been observed in individual(s) with hereditary breast and ovarian cancer (PMID: 18489799, 27062684, 29084914). This variant is also known as IVS5-2A>G. ClinVar contains an entry for this variant (Variation ID: 37923). Based on a multifactorial likelihood algorithm using genetic, in silico, and/or statistical data, this variant has been determined to have a high probability of being pathogenic (PMID: 17924331, 21990134). Studies have shown that disruption of this splice site results in skipping of exon 6 and introduces a premature termination codon (Invitae). The resulting mRNA is expected to undergo nonsense-mediated decay. For these reasons, this variant has been classified as Pathogenic. -
Pathogenic, criteria provided, single submitter	clinical testing	Women's Health and Genetics/Laboratory Corporation of America, LabCorp	Dec 05, 2022	Variant summary: BRCA2 c.476-2A>G is located in a canonical splice-site and is predicted to affect mRNA splicing resulting in a significantly altered protein due to either exon skipping, shortening, or inclusion of intronic material. Several computational tools predict a significant impact on normal splicing: Four predict the variant abolishes a 3' acceptor site. RT-PCR analysis showed the variant to result in two abnormal transcripts, one lacking exon 6 and one lacking exon 5-6, both predicted to result in premature termination codons (Colombo_2013). The variant was absent in 251060 control chromosomes. c.476-2A>G has been reported in the literature in multiple individuals affected with Hereditary Breast And Ovarian Cancer Syndrome (eg. Rebbeck_2018). These data indicate that the variant is very likely to be associated with disease. Seven clinical diagnostic laboratories have submitted clinical-significance assessments for this variant to ClinVar after 2014 without evidence for independent evaluation. All laboratories classified the variant as pathogenic. Based on the evidence outlined above, the variant was classified as pathogenic. -

Breast and/or ovarian cancer Pathogenic:1

Pathogenic, no assertion criteria provided

case-control;clinical testing

CZECANCA consortium

Jun 11, 2019

- -

Hereditary cancer-predisposing syndrome Pathogenic:1

Pathogenic, criteria provided, single submitter

clinical testing

Ambry Genetics

Dec 16, 2022

The c.476-2A>G intronic pathogenic mutation results from an A to G substitution two nucleotides before coding exon 5 of the BRCA2 gene. This mutation was found in 4 of 1010 unrelated individuals from high-risk breast and/or ovarian cancer families in the Czech Republic (Machockova E et al. BMC Cancer. 2008 May 20;8:140). RNA studies have demonstrated that this alteration results in abnormal splicing in the set of samples tested (Ambry internal data; Colombo M et al. PLoS One. 2013;8(2):e57173; Machockova E et al. BMC Cancer. 2008 May 20;8:140). This alteration is unable to complement cell growth in a BRCA2-null mouse embryonic stem cell assay (Mesman RLS et al. Genet Med, 2020 08;22:1355-1365). In addition to the clinical data presented in the literature, alterations that disrupt the canonical splice site are expected to cause aberrant splicing, resulting in an abnormal protein or a transcript that is subject to nonsense-mediated mRNA decay. As such, this alteration is classified as a disease-causing mutation. -

Familial cancer of breast Pathogenic:1

Pathogenic, criteria provided, single submitter

clinical testing

Baylor Genetics

Mar 08, 2023

- -

Computational scores

Source: dbNSFP v4.3

Name	Calibrated prediction	Score	Prediction
BayesDel_addAF	Benign	-0.14	T
BayesDel_noAF	Benign	-0.44
CADD	Pathogenic	27
DANN	Benign	0.92
Eigen	Pathogenic	0.70
Eigen_PC	Uncertain	0.44
FATHMM_MKL	Uncertain	0.91	D
MutationTaster	Benign	1.0	D;D
GERP RS		5.0

Splicing

Name	Calibrated prediction	Score	Prediction
dbscSNV1_ADA	Pathogenic	1.0
dbscSNV1_RF	Pathogenic	0.75
SpliceAI score (max)		0.98		Details are displayed if max score is > 0.2
DS_AG_spliceai		0.40		Position offset: 34
DS_AL_spliceai		0.98		Position offset: 2

Find out detailed SpliceAI scores and Pangolin per-transcript scores at spliceailookup.broadinstitute.org

Publications

LitVar

Other links and lift over

dbSNP: rs81002853; hg19: chr13-32900377;